West Nile virus (WNV) is a highly virulent human pathogen of the central nervous system (CNS) and the most common cause of epidemic encephalitis in the United States. Since there are no vaccines or specific antiviral treatments available for WNV, understanding its pathogenesis is a high priority. Following infection, WNV is transported to the local draining lymph nodes, where primary amplification of the virus and key host defense responses are triggered; however, there is very little known about the events that take place at this junction in humans. To study this, we developed an ex vivo human lymphoid tissue model of WNV infection. In preliminary studies, we show that WNV replicates robustly in all donors tested, and WNV-infected cells can be detected. In this application, we will use two strains of WNV that differ in their pathogenic potential in humans to characterize the cellular response to WNV infection and identify the permissive target cells within the lymphoid tissue (Aim 1), measure the inflammatory response during the infection (Aim 2), and determine the anatomical environment within the lymph node where virus replication preferentially occurs (Aim 3). Understanding these viral and cellular aspects in a human system and how these differ between a neuroinvasive and non-neuroinvasive strain of WNV will not only provide the first glimpse into early infection events, but also lead to new hypotheses for in vivo susceptibility in humans.